T
he essential enzymatic cofactor NAD
+ can be synt
hesized in many eukaryotes, including
Saccharomyces cerevisiae and mammals, using tryptop
han as a starting material. Metabolites along t
hepat
hway or on branc
hes
have important biological functions. For example, kynurenic acid can act as anNMDA antagonist, t
hereby functioning as a neuroprotectant in a wide range of pat
hological states.
N-Formylkynurenine formamidase (FKF) catalyzes t
he second step of t
he NAD
+ biosynt
hetic pat
hway by
hydrolyzing
N-formyl kynurenine to produce kynurenine and formate. T
he
S. cerevisiae FKF
had been reported to bea pyridoxal p
hosp
hate-dependent enzyme encoded by BNA3. We used combined crystallograp
hic,bioinformatic and bioc
hemical met
hods to demonstrate t
hat Bna3p is not an FKF but rat
her is most likelyt
he yeast kynurenine aminotransferase, w
hic
h converts kynurenine to kynurenic acid. Additionally, weidentify YDR428C, a yeast ORF coding for an
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hydrolase wit
h no previously assigned function, ast
he FKF. We predicted its function based on our interpretation of prior structural genomics results and onits sequence
homology to known FKFs. Bioc
hemical, bioinformatics, genetic and
in vivo metabolite dataderived from LC-MS demonstrate t
hat YDR428C, w
hic
h we
have designated BNA7, is t
he yeast FKF.